Antibody-induced platelet phosphatidylserine mediates increased prothrombotic potential in heparin-induced thrombocytopenia

Background

Heparin-induced thrombocytopenia (HIT) is a severe adverse event caused by antibodies (Abs) reactive to platelet factor 4 (PF4) and heparin. While HIT Abs are well characterized to activate PLTs via PLT Fc-gamma-RIIA, the contribution of different Ab-induced PLT subpopulations to the prothrombotic environment in HIT remains elusive. In this study, we hypothesized that HIT Abs have the potential to induce a procoagulant PLT phenotype that contributes to the prothrombotic condition in HIT.

Methods

Using flow cytometry (FC), we analyzed whether HIT Abs have the capability to induce changes in the expression level of the PLT surface activation marker P-selectin (CD62p) and the negatively charged procoagulant membrane phospholipid phosphatidylserine (PS). To investigate whether Ab-induced procoagulant PLTs contribute to increased thrombin generation and thrombus formation, calibrated automated thrombogram (CAT) analysis and an ex vivo thrombosis model were utilized, respectively.

Results

HIT patient Abs induced a procoagulant (CD62p/PS double positive) PLT phenotype with increased thrombin generation potential. Most importantly, spike-in of whole blood samples with HIT Ab-induced procoagulant PLTs prior to perfusion through a collagen coated microfluidic system resulted in a significant increase of thrombus formation in a heparin- as well as PLT Fc-gamma-RIIA dependent manner. Interestingly, HIT Ab-induced thrombus formation was unaffected by inhibiting the interaction of PLTs with neutrophils via CD62p-glycoprotein ligand-1 (PSGL-1) axis. Contrary, increased thrombin and most importantly, HIT Ab-induced thrombus formation were significantly reduced in the presence of the specific PS blocking protein Lactadherin.

Conclusion

Based on our mechanistic studies we conclude that increased PLT PS rather than CD62p on the surface of HIT Ab-induced procoagulant PLTs might be an essential factor for increased prothrombotic conditions typically observed in HIT.

Offenlegung Interessenkonflikt:

There a no conflicts of interest to declare.